Over the past few decades, a range of new Harmful Algal Blooms (HABs) with newly discovered natural toxins have appeared; it is thought that many of these are the result of increased nutrient loading into coastal waters and altered ecosystems particularly in subtropical and tropical marine waters. It is hypothesized that there are many other species of algae produce toxins, but these toxins go unrecognized because they are not in high enough concentrations to cause problems to humans and other animals at the present time. To understand the development of HABs, it is important to study the population dynamics of species at low concentrations before they become HABs. The central hypothesis of the proposed research project is that many species of dinoflagellates, cyanobacteria, and other algae produce toxins that are not currently noticed by humans because those species are not dense enough at the present time to produce harmful quantities of the toxins, or they are in areas of the ocean where humans do not interact much with the affected food web. As a result of increasing nutrient fluxes to the ocean, transport to new areas, global change, and changing community structure, some of these sparse or unnoticed species could generate HABs in the future. The goal of this proposed research is to examine a large number of algal species from subtropical and tropical waters for the production of a variety of types of natural marine toxins. Using the Remote Sensing Facility Core, much of the sampling regime will be guided by remote sensing data. The remote sensing data will be particularly useful in identifying "water masses" and fronts so that sampling regimes can be optimized. Temperature, salinity, oxygen, turbidity, light, and nutrients will be measured. The phytoplankton community will be characterized by pigments, flow cytometry, light microscopy, and species-specific oligonucleotide probes. Working with the Toxic Algae Culture Facility Core, individual potentially HAB species will be identified and cultured. Samples of water, sea surface microlayer, and particulates (including phytoplankton) will be analyzed for toxins by red blood cell hemolysis and ion channel activity assays. Samples that show toxicity will then be analyzed further with the other methods mentioned earlier. Using the Genomics Core, DNA sequencing will be used to determine the genetic relationships among the species, and these data will be compared to the types of toxins produced to determine possible ecological and evolutionary relationships. New species-specific oligonucleotide probes will be developed that can be used to rapidly quantify the abundance of these species in the ocean. Field sampling will be conducted primarily in subtropical South Florida coastal waters, but also in the tropical Caribbean, taking advantage of existing sampling networks used by the investigators. This sampling will examine and quantify the distribution of toxins in the sub/tropical ecosystem, the propagation of the toxins through the food web, and their spatial and temporal correlations with the species that produce the toxins. In addition, the population dynamics of non-toxic species, toxin-producing non-HAB species, and HAB species will be compared to determine if there are any fundamental differences between HAB and non-HAB species in how they respond to environmental factors and elucidate any possible ecological function for toxin production in sub/tropical marine environments.